Lead-induced cytotoxicity and transcriptional activation of stress genes in human liver carcinoma (HepG2) cells
Lead-induced cytotoxicity and transcriptional activation of stress genes in human liver carcinoma (HepG2) cells
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DOI：
10.1023/b:mcbi.0000007272.46923.12
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发表时间：
2004-01-01
影响因子：
4.3
通讯作者：
Shen, E
中科院分区：
文献类型：
--
作者：
Tchounwou, PB;Yedjou, CG;Shen, E
铅是一种非必需元素，具有高度毒性，尤其是对儿童。大多数关于铅的研究都集中在其对器官系统的影响，如神经系统、红细胞和肾脏，这些器官被认为是铅毒性的主要目标。然而，它引起毒性和致癌的分子机制仍有待阐明。在这项研究中，我们通过创建哺乳动物启动子氯霉素 (CAT) 基因融合的稳定转染子，进行了 MTT 测定来评估细胞毒性，并进行了 CAT-Tox 测定来评估与铅暴露相关的转录反应，这些重组细胞系是由人肝癌细胞 (HepG(2)) 产生的 13 种不同的重组细胞系。研究结果表明，硝酸铅对 HepG(2) 细胞具有细胞毒性，接触 24、48 和 72 小时后细胞死亡率的 LD50 值分别为 49.0 +/- 18.0 mug/mL、37.5 +/- 9.2 mug/mL 和 3.5 +/- 0.7 mug/mL；表明硝酸铅的细胞毒性作用具有剂量和时间依赖性反应。还记录了暴露于硝酸铅的 HepG(2) 细胞中应激基因诱导的剂量反应关系。总体而言，测试的 13 个重组细胞系中有 6 个显示出诱导达到统计显着水平 (p < 0.05)。在 50 μg/mL 硝酸铅下，XRE、HSP70、CRE、GADD153 和 GRP78 的平均诱导倍数分别为：2.1 +/- 1.0、5.4 +/- 0.4、12.1 +/- 6.2、5.0 +/- 1.7、2.5 +/- 1.3 和 4.8 +/- 4.5。这些结果表明硝酸铅有可能在肝脏中进行生物转化 (XRE)，导致细胞增殖 (c-fos)、蛋白质损伤 (HSP70、GRP78)、代谢扰动 (CRE) 以及生长停滞和 DNA 损伤 (GADD153)。 GSTYa (1.5 +/- 0.8) 和 GADD45 (5.7 +/- 8.1) 启动子以及 NF-kappa B (2.0 +/- 1.7) 反应元件也获得了边缘但不显着的诱导，表明氧化应激的潜力。 CYP1A1、HMTIIA、p53RE 和 RARE 没有记录到显着诱导 (p < 0.05)。
Lead is a non-essential element that exhibits a high degree of toxicity, especially in children. Most research on lead has focused on its effects on organ systems such as the nervous system, the red blood cells, and the kidneys which are considered to be the primary targets of lead toxicity. However, the molecular mechanisms by which it induces toxicity, and carcinogenesis remain to be elucidated. In this research, we performed the MTT assay to assess the cytotoxicity, and the CAT-Tox assay to assess the transcriptional responses associated with lead exposure to thirteen different recombinant cell lines generated from human liver carcinoma cells (HepG(2)), by creating stable transfectants of mammalian promoter chloramphenicol (CAT) gene fusions. Study results indicated that lead nitrate is cytotoxic to HepG(2) cells, showing LD50 values of 49.0 +/- 18.0 mug/mL, 37.5 +/- 9.2 mug/mL, and 3.5 +/- 0.7 mug/mL for cell mortality upon 24, 48 and 72 h of exposure, respectively; indicating a dose- and time-dependent response with regard to the cytotoxic effect of lead nitrate. A dose- response relationship was also recorded with respect to the induction of stress genes in HepG(2) cells exposed to lead nitrate. Overall, six out of the thirteen recombinant cell lines tested showed inductions to statistically significant levels (p < 0.05). At 50 mu g/mL of lead nitrate, the average fold inductions were: 2.1 +/- 1.0, 5.4 +/- 0.4, 12.1 +/- 6.2, 5.0 +/- 1.7, 2.5 +/- 1.3, and 4.8 +/- 4.5 for XRE, HSP70, CRE, GADD153, and GRP78, respectively. These results indicate the potential for lead nitrate to undergo biotransformation in the liver (XRE), to cause cell proliferation (c-fos), protein damage (HSP70, GRP78), metabolic perturbation (CRE), and growth arrest and DNA damage (GADD153). Marginal but not significant inductions were also obtained with the GSTYa (1.5 +/- 0.8), and GADD45 (5.7 +/- 8.1) promoters, and the NF-kappa B (2.0 +/- 1.7) response element, indicating the potential for oxidative stress. No significant inductions (p 0.05) were recorded for CYP1A1, HMTIIA, p53RE, and RARE.